In the area of PAH carcinogenesis we have performed studies using cultured normal human mammary epithelial cells (NHMECs), MCF-7 breast cancer cells, cancer-susceptible mice null for the DNA repair gene XPA and heterozygous for the tumor promoter p53, and human subjects. In chemoprevention studies using NHMECs exposed to benzo[a]pyrene (BP) we have found interindividual variability in BP-DNA adduct formation, induction of CYP1A1 and 1B1 expression, and induction of CYP1A1/1B1 enzyme (EROD) activity, all of which were reduced in the presence of chlorophyllin (CHLN) and delayed in the presence of tetramethoxystilbene (TMS). Because CHLN reduced BP-DNA damage in all of the 19 NHMEC strains studied, it appears this compound may be an appropriate chemoproventive agent. In contrast, TMS delayed BP metabolism but did not reduce the amount of BP-DNA damage generated during 96 hr of exposure. Therefore TMS does not appear to be an effective chemopreventive agent. In additional cell culture studies we examined expression of CYP1A1 and CYP1B1 as gene abundance (copy levels/ng RNA) in normal human mammary epithelial cells (NHMECs) and MCF-7 cells exposed to BP, in the presence or absence of CHLN. Endogenous and BP-induced gene abundance of CYP1B1 was 5- to 10-fold higher in MCF-7 cells compared to NHMECs. This correlated with the finding that MCF-7 cells form 10-fold more BP-DNA adducts than NHMECs. In addition, CHLN treatment did not reduce BPdG adduct levels in the MCF-7 cells. Subsequently enzyme activities of CYP1A1/CYP1B1, and the detoxification enzyme NQO1, were measured in BP-exposed NHMECs and MCF-7 cells. CYP1A1/1B1 enzyme activity, determined by EROD assay, was 4 fold higher in MCF-7 cells than in NHMEC strain 98016, and increased by 5-fold and 1.6-fold, respectively, after exposure to BP. NQO1 enzyme specific activity was 3-fold lower in MCF-7 cells than in NHMECs, and decreased slightly in both cell types after exposure to BP. Therefore, the high BPdG level observed in MCF-7 cells appears to be driven by high levels of CYP1B1 enzyme, and low levels of NQO1 enzyme. The data suggest that, compared to MCF-7 cells, NHMECs may be protected from BP-DNA damage, and possibly mutagenesis, by a high capacity for detoxification and comparatively low levels of activation. A short latency for BP-induced tumors has been shown in (XPA -/-, p53 +/-) C57BL/6 mice lacking the first step in nucleotide excision repair (NER) and haploinsufficient for p53. Mice were fed diet containing no additions, 100 ppm BP, 100 ppm BP plus 0.3% CHLN, or 0.3% CHLN alone for 28 days. The two methods used to measure BP-induced DNA damage in the esophagus, lungs and liver were: high pressure liquid chromatography tandem mass spectrometry (HPLC-MS/MS), which measures only the BPdG adduct;and, BPDE-DNA CIA using antiserum elicited against BPDE-modified DNA, which, in addition to BPdG, measures all the other stable BP-induced DNA adducts. Both methods showed that in all three organs of mice fed BP alone, the DNA adduct levels at 28 days were higher in the cancer-susceptible Xpa(-/-)p53(+/-) mice, compared to the WT mice. This observation was consistent with the relative differences in BP-induced tumor susceptibility reported previously. In addition, a comparison of methods showed that lung and liver formed more adducts detected by CIA than HPLC-MS/MS, suggesting that these organs formed many more BP-induced adducts than just BPdG. In contrast, the majority of adducts found in esophagus appeared to be BPdG. Based on many prior studies we hypothesized that addition of the chemopreventive agent CHLN to BP in the diet would reduce the level of BP-induced DNA damage in all mouse organs. In the Xpa(-/-) p53(+/-) mice fed BP plus CHLN there was evidence for a reduction in BP-DNA adduct formation in the liver, but not in the other organs. In the WT mice fed BP plus CHLN, BP-DNA adducts were increased several-fold in esophagus and liver, contrary to our expectations. The data suggest that administration of dietary CHLN, along with BP, may not result in reduced BP-induced DNA damage in all organs, and therefore use of CHLN as a chemopreventive agent in the human population should be approached with caution. Parental exposure to tobacco smoke carcinogens has been implicated in the development of childhood cancers. Cytotrophoblast (CT) cells and syncytiotrophoblast (ST) knots that line the placental chorionic villi are known to metabolize xenobiotics. In this study we examined human placenta for PAH-DNA adduct formation using semi-quantitative immunohistochemistry (IHC) with Automated Cellular Imaging System (ACIS) analysis, and a standard curve of cultured human keratinocytes exposed for 1 hr to increasing doses of r7, t8-dihydroxy-t-9, 10-oxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE). Standard curve cells were evaluated both by IHC/ACIS, and by BPDE-DNA chemiluminescence immunoassay (CIA) of extracted DNA, and the two approaches were highly correlated. Full-term human placenta samples showed positive nuclear staining localized in CT cells and ST knots. Comparison of placental IHC values with the keratinocyte standard curve revealed statistically indistinguishable PAH-DNA adduct values for 7 smokers and 7 non-smokers. The data confirm the efficiency of CT cells and ST knots for metabolism of PAHs, and suggest that PAH-DNA adduct formation in human placenta occurs as a result of multiple types of exposures. In an additional study, 184 non-smoking young adults living in Mexico City were shown to have higher levels of PAH-DNA adducts in the dry season (winter-spring), compared to the rainy season (summer-autumn). PAH-DNA adducts correlated with the seasonal levels of pollution as measured by chemcials extracted from PM10 filters, so that levels of pollution and PAH-DNA adducts were significantly higher (p&lt;0.001) in the dry season than in the rainy season. The formation of TAM-DNA adducts in human endometrium is a controversial topic of interest, as TAM-exposed women are at risk for endometrial cancer. We have examined this question in multiple tissues of aging Erythrocebus patas (patas) monkeys given oral TAM dosing, as well as in human endometrial biopsy samples. Adducts were determined by TAM-DNA chemiluminescence immuno-assay (CIA) in 3 female patas monkeys given oral TAM dosing for 3 months and 2 unexposed controls. The highest TAM-DNA adduct levels were found in liver, uterus, and brain, and none were detectable in ovary and kidney. In humans, samples of normal endometrium or endometrial tumor from patients receiving TAM therapy (n=8) had measurable TAM-DNA values, while no TAM-DNA adducts were detected in 8 patients not receiving TAM therapy. Therefore, TAM-DNA adducts are formed in primate reproductive organ tissues. Interestingly, the TAM-DNA adduct levels in uterus of patas monkeys given a daily TAM dose 5-fold higher than the human daily TAM dose, were about 3-fold higher than the TAM-DNA adduct levels found in human endometrium. The data suggest that TAM-DNA adducts can be formed in endometrium of human patients and that DNA damage may contribute to the risk of endometrial cancer.